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In cell respiration, the proton pump uses energy to transport protons from the matrix of the mitochondrion to the inter-membrane space.[1] It is an active pump, that generates a proton concentration gradient across the inner mitochondrial membrane, because there are more protons outside the matrix than inside. The difference in pH and electric charge (ignoring differences in buffer capacity) creates an electrochemical potential difference that works similar to that of a battery or energy storing unit for the cell.[2] The process could also be seen as analogous to cycling uphill or charging a battery for later use, as it produces potential energy. The proton pump does not create energy, but forms a gradient that stores energy for later use.

The FoF1ATP synthase of mitochondria, in contrast, usually conduct protons from high to low concentration across the membrane while drawing energy from this flow to synthesize ATP. Protons translocate across the inner mitochondrial membrane via proton wire. This series of conformational changes, channeled through the a and b subunits of the F0 subunit, drives a series of conformational changes in the stalk connecting the F0 to the F1 subunit. This process effectively couples the translocation of protons to the mechanical motion between the Loose, Tight, and Open states of F1 necessary to phosphorylate ADP.

In addition to proton pumps in mitochondria, humans (and probably other mammals) have a gastric hydrogen potassium ATPase or H+/K+ ATPase that functions as the proton pump of the stomach, primarily responsible for the acidification of the stomach contents (see gastric acid).